Preimplantation genetic testing for aneuploidy and chromosomal structural rearrangement: A summary of a nationwide study by the Japan Society of Obstetrics and Gynecology

Abstract Purpose The Japan Society of Obstetrics and Gynecology conducted a nationwide clinical study to evaluate the pregnancy outcomes of preimplantation genetic testing for aneuploidy or chromosomal structural rearrangement (PGT‐A/SR). Methods Patients that had experienced recurrent implantation failure, recurrent pregnancy loss, or chromosomal structural rearrangement were recruited from 200 fertility centers in Japan. For patients in whom one or more blastocysts were classified as euploid or euploid with suspected mosaicism, a frozen–thawed single embryo transfer (ET) was performed. Results A total of 10 602 cycles, maternal age 28–50 years, were enrolled in this study. 42 529 blastocysts were biopsied, and 25.5%, 11.7%, and 61.7% of embryos exhibited euploidy, mosaicism, and aneuploidy, respectively. At least one euploid blastocyst was obtained in 38.3% of egg retrieval cycles with embryo biopsy. A total of 6080 ETs were carried out, and the clinical pregnancy rate per ET, ongoing pregnancy rate per ET, and miscarriage rate per pregnancy were 68.8%, 56.3%, and 10.4%, respectively. The rates of clinical pregnancy and miscarriage remained relatively constant across all maternal ages. Conclusions Preimplantation genetic testing for aneuploidy or chromosomal structural rearrangement may improve the pregnancy rate per ET and reduce the miscarriage rate per pregnancy, especially in patients of advanced maternal age.


| INTRODUC TI ON
Aneuploidy in gametes and embryos is a major cause of implantation failure during in vitro fertilization (IVF) and miscarriage. 1-3 As most aneuploidies arise in maternal meiosis, and they are more common in older women, 4 the reproductive outcomes of IVF are worse in patients of advanced maternal age. In 2020, the Japan Society of Obstetrics and Gynecology (JSOG) annual online cycle-based assisted reproductive technology (ART) registry showed that the pregnancy rate and live birth rate per embryo transfer (ET) were 15.8% and 9.9% or lower, and the miscarriage rate per pregnancy was 33.3% or higher in women of 40 years of age or older. 5 In addition, chromosomal structural rearrangements (CRs), including Robertsonian translocations, reciprocal or balanced translocations, and inversions, are well-known risk factors for miscarriage. Although morphological assessments are the primary method for embryo prioritization, neither static nor dynamic evaluations can accurately determine chromosome status.
Preimplantation genetic testing for aneuploidy or chromosomal structural rearrangement (PGT-A/SR) is based on methods for selecting embryos with high potential for implantation and pregnancy and low risk of miscarriage. 6,7 Initially, cleavage-stage biopsies and fluorescence in situ hybridization (FISH) were used for PGT-A/SR, but their efficacy could not be confirmed in randomized control trials (RCT). 8 Recently, molecular techniques, blastocyst culturing, and embryo vitrification have improved. As a result, they have been extensively utilized for PGT-A, and some studies have shown that PGT-A produces favorable pregnancy outcomes per ET in limited infertile populations. [9][10][11][12] On the other hand, several studies have failed to demonstrate beneficial effects of PGT-A, even after the improvement of PGT-A techniques, especially in young populations, [13][14][15] and it remains unclear whether PGT-A increases the cumulative live birth rate per egg retrieval cycle or intention to treat. 16 In addition, because PGT-A, but not PGT-SR, was prohibited in Japan for a long time, the effects of PGT-A on infertile Japanese patients have not been elucidated. 17 A previous pilot study conducted by the Japan Society of Obstetrics and Gynecology (JSOG) showed that PGT-A improved the live birth rate per ET in patients that had experienced recurrent implantation failure (RIF) or recurrent pregnancy loss (RPL), but did not improve the live birth rate per patient or reduce the miscarriage rate. 18 However, it is possible that the sample size of this pilot study was too small to detect a significant beneficial effect on the clinical miscarriage rate. Therefore, JSOG conducted a nationwide clinical study with a large study population to evaluate the pregnancy outcomes of PGT-A/SR in patients that had experienced RIF or RPL or exhibited CR. Here, we summarize the data for 10 602 registered cycles collected in this clinical study.

| Study design
We conducted a multi-center open-label clinical trial, involving patients recruited from 200 fertility centers and testing at 17 laboratories in Japan. Each fertility center followed their own standard of care regarding ovarian stimulation, oocyte retrieval, IVF procedures, endometrial preparation, luteal-phase support, and ET. In each case, a trophectoderm (TE) biopsy was performed on a good quality blastocyst from around five TE cells located apart from the inner cell mass, and the biopsy sample was then transferred to a genetic testing laboratory. After the TE biopsy, the blastocysts were vitrified.
The genetic testing laboratories were required to have established processes based on their own internally validated testing/ reports and to meet known sequencing quality metrics. This study was approved by the research ethics committee of the JSOG and Tokushima University Hospital. In addition, appropriate approvals were obtained from institutional review boards or ethics committees in a site-specific manner. Written informed consent was obtained from each couple before the procedures were performed. Patients that exhibited CR during IVF-ET were also enrolled regardless of whether they had a history of pregnancy and miscarriage. The only exclusion criterion for CR was severe complications. Enrollment was completed before oocyte retrieval, and the patients received no financial incentives for participation. All cycles were registered temporarily, and full registration was permitted after the study protocol had been completed. Cycles in which full registration did not occur within 6 months of temporary registration were regarded as dropouts.

| Outcomes
The primary study outcome was the ongoing pregnancy rate at 12 weeks of gestation for each enrolled patient. The secondary study outcomes were the clinical pregnancy rate per ET and the miscarriages rate per clinical pregnancy. Cases in which a gestational sac formed were diagnosed as clinical pregnancies, and cases in which spontaneous or unplanned loss of a fetus from the uterus before 12 weeks of gestation were diagnosed as miscarriage. Cases with missing outcomes were excluded from analysis.

| RE SULTS
During the study period, a total of 10 602 cycles (maternal age 28-

| DISCUSS ION
In this study, we summarized the data collected in a nationwide clinical study of PGT-A/SR conducted by JSOG. During the study period, of the cases. Regarding the distribution of maternal age, the mean age of the patients that underwent PGT-A/SR was 39.1 years of age; a little older than the mean age (37.8 years of age) of the patients that underwent ART in Japan in 2020. 5 As around 90% of patients had undergone several cycles of ART, and some of them had experienced two or more miscarriages before participating in this study, the higher maternal age of the patients that underwent PGT-A/SR in this study may have been due to their longer treatment history.
In total, the euploidy rate was 25.5% in this study, and at least one euploid blastocyst was obtained in 38.3% of egg retrieval cycles. These results are similar to those obtained in our pilot study, 18 whereas they are much lower than those seen in previous studies performed in other countries. 4,19 Euploid blastocysts and euploid blastocysts with suspected mosaicism were classified into separate categories in this study, whereas these two groups were simply categorized as euploid blastocysts in previous studies and the criteria for designating mosaicism differ among centers. 20 These differences in definitions may explain the discrepancies in the abovementioned results. The proportion of euploid embryos decreased after 37 years of age and reached <15% at 43 years of age or older. This tendency is similar to those seen in previous studies, for example, the euploid rate was highest between the ages of 26 and 30 and steadily decreased through to age 43 and then plateaued. 4 Taken together, these findings indicate that around 40% of patients may produce at least one euploid blastocyst per egg retrieval cycle and that the euploid blastocyst per egg retrieval cycle rate decreases as maternal In this study, the clinical pregnancy rate per ET was about 68.8%, and the miscarriage rate per pregnancy was about 10.4%, and these pregnancy and miscarriage rates continued to be seen at advanced maternal age. In contrast, it has been reported that the pregnancy rate per ET was 33.9% and the miscarriage rate per pregnancy was 24.9% in patients that underwent ART in Japan in 2020 and that these rates worsened with age. 5

ACK N OWLED G M ENTS
The authors wish to thank all the subcommittee members of JSOG and the facilities that participated in this JSOG study.

CO N FLI C T O F I NTE R E S T S TATE M E NT
The authors declare no conflict of interest.

E TH I C S S TATEM ENT
This study was approved by the research ethics committee of the JSOG and Tokushima University Hospital. In addition, site-specific appropriate institutional review board or ethics committee approval was obtained.

H U M A N R I G HT S S TATE M E NT S A N D I N FO R M E D CO N S E NT
All procedures were performed in accordance with the ethical standards of the relevant committees on human experimentation (institutional and national) and the Helsinki Declaration of 1964 and its later amendments.

A N I M A L R I G HTS
This report does not contain any studies performed by any authors that included animals.